During folic acid and vitamin B12 deficiencies, hyperhomocystenemia occurs due to impaired methylation of homocysteine (Hcy) to form methionine. Hcy is the immediate precursor of the amino acid, methionine. In humans, Hcy is of great interest because it has been identified as a putative risk factor for several age-related disease states including arteriosclerosis, myocardial infarction, peripheral arterial occlusive disease, subcortical vascular encephelopathy, Alzheimer's disease and neural tube defects. Increased plasma Hcy has also been found in patients with AIDS, arthritis, and multiple sclerosis. Folic acid and vitamin B12 deficiency have also been independently associated with decreased immune function, the apoptosis of bone marrow hematopoietic progenitor cells and the appearance of leukocytes with hypomethylated DNA in the peripheral circulation. Mechanisms for these observations have been described almost exclusively in terms of the obligate role for folic acid and vitamin B12 in the methylation of genomic DNA. A specific role for Hcy or its metabolites in these processes has not been described. Our initial results reveal that treatment of resting human T cells with the combination of serum-free media and Hcy (250 uM to 1000 uM) resulted in a dose-dependent increase in apoptotic cell death. Hcy was more potent than Hcy thiolactone in this respect while S-adenosyl Hcy was inactive. This Hcy-induced T cell death could not be prevented by pretreatment with either folic acid or cycloheximide. However, this effect was either partially or fully inhibited by pretreatment with human serum, caspase inhibitors and poly-ADP-ribose polymerase inhibitors. In addition, Hcy-induced apoptosis was abrogated by pretreatment of T cells by orthovanadate, intracellular calcium chelation or cellular activation via anti-CD3/CD28 antibodies. Hcy (250 uM to 1000 uM) also potentiated T cell death induced by heat shock but not by gamma-irradiation (140 to 1400 rad) in serum-containing media. Finally, we have also found that Hcy induces Th1 cytokine production within these cultures and facilitates enhanced actvation-induced cell death of these cells. Together, these results suggest a possible role for Hcy as a pro-apototic stimulator of human T cells.